Summary Approximately 20-30% of patients with metastatic breast cancer develop brain metastasis, which is almost always fatal, and the median survival time of these patients is only few months. This observation is particularly imminent for patients with triple negative tumors. Ironically, recent advancement in chemotherapy has further increased the incidence of brain metastasis because most therapeutic agents cannot effectively penetrate the BBB and tumor cells find the brain as a sanctuary. Therefore, it is of paramount importance to elucidate the molecular mechanism involved in the metastatic process in order to define a specific therapeutic target. The recent cancer stem cell theory predicts that metastasis initiating cells (MIC) are by definition cancer stem-like cells (CSC). However, how MICs adapt themselves or passively generate a niche in the brain is a critical question. We have recently found that (i) brain MICs highly express the key inflammatory cytokine, IL-1?, which activates astrocytes, (ii) the secreted IL-1? up-regulates Jagged1, a Notch ligand, in the reactivated astrocytes, which in turn activates Notch signaling in MICs, (iii) IL-1? also mobilizes a microRNA network by activating miR301 and miR629, (iv) miR301 and miR629 are secreted from cancer cells as exosomes and suppress VHL expression, which results in HIF-1 and VEGF-D activation in astrocytes, and (v) VEGF-D then promotes MIC self-renewal. Based on these preliminary data, we hypothesize that brain MICs secrete IL-1?, which up-regulates Notch ligand expression in astrocytes and also stimulates the release of exosomal miR301/miR629 from the tumor cells that together activate cancer associated astrocytes, thereby, promoting MIC self-renewal. We also hypothesize that the secreted exosomes can serve as biomarkers for brain metastasis at an early stage and that a BBB-permeable and thermo-activatable pro-drugs can suppress brain metastasis progression by blocking IL-1? and Notch signaling. To test these hypotheses, we will first elucidate the roles of exosomal microRNAs in the metastatic niche generation in the brain (Aim 1). We will also define the role of IL1?-mediated activation of a microRNA network and Notch pathway in self-renewal of MICs in brain metastasis (Aim 2). We will then examine the efficacy of a BBB-permeable and thermo-activatable pro-drug on brain metastasis (Aim 3). Our study is highly innovative in that (i) we have identified the critical role of the brain microenvironment, particularly astrocytes, in the progression of MIC self- renewal, (ii) we showed that IL-1? also promote the secretion of exosomal microRNAs (miR301/miR629), which act as messengers for cell-cell communications in the stem cell niche, (iii) we will explore whether these exosomal microRNAs can be used as biomarkers for early diagnosis of brain metastasis, (iv) we will develop an innovative IL1RA pro-drug to target both tumor and microenvironmental cells in the brain. We believe that the outcome of our study will provide a paradigm shift in our current understanding of the pathology of brain metastasis and also have a significant impact on early detection of brain metastasis as well as on future treatments for this devastating disease.